The project involves the immunosuppressive agents cyclosporin A, FK506 and rapamycin; their interactions with their binding proteins cyclophilin A (CypA) and FKBP12; the interaction of the binary complexes with the ultimate cellular targets such as FRAP; and finally the generalization of these studies into the design of small cell-permeable agents that can dimerize proteins. Most of our work in the last period has emphasized the structure of FKBP12-rapamycin-FRB and the design of 'hole-bump'pairs based on CypA - CysA. We were able to collect a mediocre, 2.8 _ resolution, data set on the FKBP12-rapamycin-FRB on a laboratory instrument. Using MacCHESS resources (F1, CCD), we've collected a 2.0 _ resolution data set and are just finishing our analysis of it. We have also used F1 to study the uncomplexed FRB domain. This analysis is not yet complete, but it could provide crucial support for the hypothesis that the FRB domain is an ligand controlled conformational switch for controlling the kinase activity of FRAP and homologs. In order to begin a structure-based drug design project on dimerizing agents for controlling cellular processes, a cyclosporin analog, CysA11MeIle was synthesized. This mutation abolishes binding to wild type CypA (Kd > 3 uM). A double mutant of Cyp A (S99T, F113A) was found to bind the mutated CysA tightly (Kd = 2 nM). We crystallized the complex in order to see structural alterations responsible for this tight binding, but the crystals (P61) had a long axisrepeat--greater than 400 _. In January, we were able to collect a data set on F1 using the CCD detector. These data were good to 3.5 _ and yielded a MR solution. In May, another data set was collected on F1 using Fuji image places, and these data extent to 2.8 _ and are now being analyzed.